209 results on '"Costa, R. H."'
Search Results
52. Herpes simplex virus mRNA species mapping in EcoRI fragment I
- Author
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Hall, L M, Draper, K G, Frink, R J, Costa, R H, and Wagner, E K
- Abstract
We described the detailed characterization and high-resolution mapping of nine herpes simplex virus type 1 mRNAs encoded in EcoRI fragment I. Four of these mRNAs are partially colinear and encode the same sized polypeptide in vitro. Nucleotide sequence analysis of the DNA around the 5' ends of these mRNAs suggested that the larger may encode a small (ca. 100-dalton) polypeptide not resolvable by in vitro translation.
- Published
- 1982
- Full Text
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53. A liver-specific DNA-binding protein recognizes multiple nucleotide sites in regulatory regions of transthyretin, alpha 1-antitrypsin, albumin, and simian virus 40 genes.
- Author
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Costa, R H, Grayson, D R, Xanthopoulos, K G, and Darnell, J E
- Abstract
Double-stranded oligodeoxynucleotides that represent protein binding sites in the regulatory regions of the mouse genes encoding transthyretin (TTR) and alpha 1-antitrypsin (alpha 1-AT) bound a nuclear protein factor(s) found mainly in hepatocytes. A site in the regulatory region of the gene encoding rat serum albumin and, surprisingly, a region in the simian virus 40 enhancer also bind the same factor. Oligodeoxynucleotide affinity chromatography (with one of the TTR binding sites) allowed a 500-fold purification of the protein. The purified protein protected similar portions of all the regulatory regions, as well as the simian virus 40 core C enhancer element, from digestion with DNase I. A DNA-binding protein previously purified from liver by virtue of its ability to bind to several virus enhancer sequences also binds to TTR, alpha 1-AT, and albumin regulatory sites. Thus, all these binding sites, which contain only minimal sequence similarity, may bind to a single protein, or a similar family of proteins, that activates liver-specific transcription of coordinately expressed genes.
- Published
- 1988
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54. The cell-specific enhancer of the mouse transthyretin (prealbumin) gene binds a common factor at one site and a liver-specific factor(s) at two other sites
- Author
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Costa, R H, Lai, E, Grayson, D R, and Darnell, J E
- Abstract
We previously defined two distinct cell-specific DNA elements controlling the transient expression of the transthyretin gene in Hep G2 (human hepatoma) cells: a proximal promoter region (-202 base pairs [bp] to the cap site), and a far-upstream cell-specific enhancer located between 1.6 and 2.15 kilobases (kb) 5' of the cap site (R. H. Costa, E. Lai, and J. E. Darnell, Jr., Mol. Cell. Biol. 6:4697-4708, 1986). In this report, we located the effective transthyretin enhancer element within a 100-bp region between 1.96 and 1.86 kb 5' to the mRNA cap site. In Hep G2 nuclear extracts, three protein-binding sites within this minimal enhancer element were identified by gel mobility and methylation protection experiments. Each binding site was required for full enhancer activity in Hep G2 transient expression assays. Competition experiments in protein-binding assays suggested that two of the three sites were recognized by a similar factor and that the protein interaction with the third site was different. The nuclear protein(s) which bound to the two homologous sites was found mainly or only in cells of hepatic origin, suggesting an involvement of this region in the cell-specific function of this enhancer. The nuclear protein(s) recognizing the third enhancer region was also found in HeLa and spleen cells.
- Published
- 1988
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55. Characterization of the genes encoding herpes simplex virus type 1 and type 2 alkaline exonucleases and overlapping proteins
- Author
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Draper, K G, Devi-Rao, G, Costa, R H, Blair, E D, Thompson, R L, and Wagner, E K
- Abstract
A detailed sequence analysis of the herpes simplex virus type 1 (HSV-1) and HSV-2 DNA encoding the alkaline exonuclease mRNA clusters has been completed. Three partially colinear mRNAs (2.3, 1.9, and 0.9 kilobases) are completely encoded within the DNA sequence presented. The putative promoter regions of the transcripts were inserted upstream of a plasmid-borne chloramphenicol acetyl transferase (CAT) gene and assayed for their ability to induce transcription of the CAT gene upon low multiplicity of infection with HSV in transient expression assays. We conclude that the expression of all three transcripts appear to be controlled by individual promoters. The 2.3-kilobase mRNA contains an open translational reading frame sufficient to encode 626 amino acids for the HSV-1 alkaline exonuclease enzyme; this value is 620 amino acids for HSV-2. A comparison of the predicted amino acid sequences of the HSV-1 and HSV-2 alkaline exonuclease enzymes revealed significant amino acid differences in the N-terminal portions of the two proteins; however, computer analyses suggest that the three-dimensional structures of the HSV-1 and HSV-2 nuclease enzymes are very similar. The 0.9-kilobase mRNA contains an open reading frame which shares a small amount of out-of-phase overlap with the C-terminal portion of the alkaline nuclease open reading frame. This open reading frame has the capacity to encode a 96-amino-acid polypeptide (10,500 daltons).
- Published
- 1986
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56. The transcriptional activator hepatocyte nuclear factor 6 regulates liver gene expression
- Author
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Samadani, U and Costa, R H
- Abstract
The hepatocyte nuclear factor 3(alpha) (HNF-3(alpha)), -3(beta), and -3(gamma) proteins share homology in the winged-helix/fork head DNA binding domain and mediate hepatocyte-enriched transcription of numerous genes whose expression is necessary for organ function. In this work, we identify a liver-enriched transcription factor, HNF-6, which recognizes the -138 to -126 region of the HNF-3(beta) promoter and binds the original HNF-3 site of the transthyretin promoter (-94 to -106). We show that HNF-6 and HNF-3 possess different DNA binding specificities by competition and methylation interference studies and are immunologically distinct. Site-directed mutagenesis of the HNF-6 sites in the HNF-3(beta) and transthyretin promoters diminishes reporter gene expression, suggesting that HNF-6 activates transcription of these promoters. Using the HNF-6 binding sequence DHWATTGAYTWWD (where W = A or T, Y = T or C, H is not G, and D is not C) determined by sequence comparison and methylation interference, we predicted that HNF-6 will bind to 22 additional hepatocyte-enriched genes. Of these potential target genes, we selected seven of the HNF-6 binding sequences and demonstrated that they bind the HNF-6 protein. These include promoter sequences from alpha-2 urinary globulin, alpha-1 antitrypsin, cytochrome P-450 2C13, L-type 6-phosphofructo-2-kinase, mouse major urinary protein, tryptophan oxygenase, and alpha-fetoprotein genes. HNF-6 binding activity was also found in the intestinal epithelial cell line HT29, and potential HNF-6 binding sites were present in intestinal sucrase isomaltase, cdx-2 homeodomain protein, and intestinal fatty acid binding protein promoter regions. These studies suggest that HNF-6 may regulate hepatocyte-specific genes and may play a role in epithelial cell differentiation of gut endoderm via regulation of HNF-3(beta).
- Published
- 1996
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57. An unusual spliced herpes simplex virus type 1 transcript with sequence homology to Epstein-Barr virus DNA
- Author
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Costa, R H, Draper, K G, Kelly, T J, and Wagner, E K
- Abstract
High-resolution transcription mapping localized a spliced 2.7-kilobase herpes simplex virus type 1 mRNA. The 4-kilobase intron of this transcript encodes a nested set of transcripts on the opposite DNA strand. The nucleotide sequence of the DNA encoding the left-hand and right-hand exons of the spliced transcript was determined, and the salient features are presented here. Of major interest is that both exons contained regions within several hundred bases of the splice donor and acceptor sites which showed homology to two regions of the Epstein-Barr virus genome, which are themselves 3 kilobases apart. The spliced herpes simplex virus transcript encoded a translational reading frame which could encode a protein with an approximate size of 75,000 daltons. This value is in agreement with in vitro translation data. The predicted amino acid sequence of the herpes simplex virus protein had significant homology with putative amino acid sequences encoded by the homologous Epstein-Barr virus DNA sequences.
- Published
- 1985
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58. Multiple hepatocyte-enriched nuclear factors function in the regulation of transthyretin and alpha 1-antitrypsin genes
- Author
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Costa, R H, Grayson, D R, and Darnell, J E
- Abstract
Transthyretin (TTR) and alpha 1-antitrypsin (alpha 1-AT) are expressed at high levels in the liver and also in at least one other cell type. We report here a detailed analysis of the proximal regulatory region of the TTR gene, which has uncovered two new DNA-binding factors that are present mainly (or only) in hepatocytes. One of these new factors, hepatocyte nuclear factor 3 (HNF-3), binds to two sites that are crucial in TTR expression as well as to two additional sites in the alpha 1-AT proximal enhancer region. The second new factor, HNF-4, binds to two sites in TTR that are required for gene activity. We had previously identified binding sites for another hepatocyte-enriched DNA-binding protein (C/EBP or a relative thereof), and additional promoter-proximal sites for that protein in both TTR and alpha 1-AT are also reported here. From these results it seems clear that cell-specific expression is not simply the result of a single cell-specific factor for each gene but the result of a combination of such factors. The variation and distribution of such factors among different cell types could be an important basis for the coordinate expression of the TTR and alpha 1-AT genes in the liver or the discordant transcriptional activation of these genes in a few other cell types. The identification of such cell-enriched factors is a necessary prelude to understanding the basis for cell specificity.
- Published
- 1989
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59. Performance of a baffled facultative pond treating piggery wastes
- Author
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Zanotelli, C. T., Medri, W., Paulo Belli Filho, Perdomo, C. C., Mulinari, M. R., and Costa, R. H. R.
60. Reduction of odors from a facultative pond using two different operating practices
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Truppel, A., Camargos, J. L. M., Da Costa, R. H. R., and Paulo Belli Filho
61. The Epistolary Monitor in Pamela
- Author
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Costa, R. H., primary
- Published
- 1970
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62. High-rate pond for treatment of piggery wastes
- Author
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Medri, W., Costa, R. H. R., and Perdomo, C. C.
- Subjects
- *
LIVESTOCK , *SEWAGE , *SEWAGE purification , *HYDROLOGY - Abstract
This work deals with studies on high-rate ponds, a batch working system that is followed by a filter pond where Chinese carp were introduced for piggery wastes treatment. COD removal values for the high-rate pond were in the order of 95% in the summer and 70% in the winter for an initial concentration of 2000 mg/L. Total nitrogen removal values ranged between 90% and 60%, respectively, in summer and winter periods for an initial concentration of 600 mg/L. Seasonal variations, which are mainly observed under differences of temperature, were shownto be not relevant for total phosphorus removal, a process that appears to depend mainly on increases of pH values over 8.5. An hydraulicretention time ranging between 15 to 20 days was found to be best for pond functioning. The plug flow model fits well to the pond's physical characteristics. The filter pond was shown to be a great potential process for removal of algae produced in the high-rate pond. [ABSTRACT FROM AUTHOR]
- Published
- 2000
63. Tertiary treatment of piggery wastes in water hyacinth ponds
- Author
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Bavaresco, A. S. L., Philippi, L. S., Medri, W., and Costa, R. H. R.
- Subjects
SEWAGE purification ,SEWAGE ,HYDROLOGY ,LIVESTOCK - Abstract
This work was developed in two stages: initially in water hyacinth ponds under batch conditions (pilot-scale) and later in full-scale ponds under a continuous state, so as to determine local parameters for the operation of this kind of pond for piggery wastes treatment. Seasonal variations are important features influencing plant productivityand performance. The water hyacinth ponds were able to remove around50% of the applied organic loads (COD, BOD, TN, TP), even in case ofelevated values (110 kg/ha/day) applied for total nitrogen surface loads. An hydraulic retention time of 20 days was shown to be ideal for the treatment. The utilization of water hyacinths as a complement for the animal diet closes the productive cycle in a sustainable way. The results have demonstrated the feasibility of the proposed treatment system. [ABSTRACT FROM AUTHOR]
- Published
- 2000
64. Effect of COD/N ratio on N 2 O production during nitrogen removal by aerobic granular sludge.
- Author
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Velho VF, Magnus BS, Daudt GC, Xavier JA, Guimarães LB, and Costa RHR
- Subjects
- Aerobiosis, Ammonia, Biological Oxygen Demand Analysis, Denitrification, Heterotrophic Processes, Nitrification, Nitrites, Nitrogen metabolism, Nitrous Oxide analysis, Waste Disposal, Fluid methods, Bioreactors microbiology, Nitrogen chemistry, Nitrous Oxide metabolism, Sewage microbiology
- Abstract
N
2 O-production was investigated during nitrogen removal using aerobic granular sludge (AGS) technology. A pilot sequencing batch reactor (SBR) with AGS achieved an effluent in accordance with national discharge limits, although presented a nitrite accumulation rate of 95.79% with no simultaneous nitrification-denitrification. N2 O production was 2.06 mg L-1 during the anoxic phase, with N2 O emission during air pulses and the aeration phase of 1.6% of the nitrogen loading rate. Batch tests with AGS from the pilot reactor verified that at the greatest COD/N ratio (1.55), the N2 O production (1.08 mgN2 O-N L-1 ) and consumption (up to 0.05 mgN2 O-N L-1 ), resulted in the lowest remaining dissolved N2 O (0.03 mgN2 O-N L-1 ), stripping the minimum N2 O gas (0.018 mgN2 O-N L-1 ). Conversely, the carbon supply shortage, under low C/N ratios, increased N2 O emission (0.040 mgN2 O-N L-1 ), due to incomplete denitrification. High abundance of ammonia-oxidizing and low abundance of nitrite-oxidizing bacteria were found, corroborating the fact of partial nitrification. A denitrifying heterotrophic community, represented mainly by Pseudoxanthomonas, was predominant in the AGS. Overall, the AGS showed stable partial nitrification ability representing capital and operating cost savings. The SBR operation flexibility could be advantageous for controlling N2 O emissions, and extending the anoxic phase would benefit complete denitrification in cases of low C/N influents.- Published
- 2017
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65. Anaerobic side-stream reactor for excess sludge reduction: 5-year management of a full-scale plant.
- Author
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Velho VF, Foladori P, Andreottola G, and Costa RH
- Subjects
- Aerobiosis, Anaerobiosis, Bioreactors, Sewage, Waste Management methods, Wastewater
- Abstract
The long-term performances of a full-scale anaerobic side-stream reactor (ASSR) aimed at sludge reduction have been monitored for the first time, in comparison with a conventional activated sludge process (CAS). The plant was integrated with an ASSR treatment of 2293-3293 m(3). Operational parameters in the ASSR were: ORP -250 mV, interchange ratio of 7-10%, hydraulic retention time of 7 d. No worsening of effluent quality was observed in the ASSR configuration and removal efficiency of COD and NH4 was above 95%. A slight increase in the Sludge Volume Index did not cause worsening in effluent solids concentration. The observed sludge yield (Yobs) passed from 0.44 kgTSS/kgCOD in the CAS to 0.35 in the ASSR configuration. The reduction of Yobs by 20% is lower than expected from the literature where sythetic wastewater is used, indicating that sludge reduction efficiency is largely affected by inert mass fed with influent real wastewater. An increase by 45% of the ASSR volume did not promote a further reduction of Yobs, because sludge reduction is affected not solely by endogenous decay but also by other factors such as interchange ratio and aerobiosis/anaerobiosis alternation., (Copyright © 2016 Elsevier Ltd. All rights reserved.)
- Published
- 2016
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66. Concerning the role of cell lysis-cryptic growth in anaerobic side-stream reactors: the single-cell analysis of viable, dead and lysed bacteria.
- Author
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Foladori P, Velho VF, Costa RH, Bruni L, Quaranta A, and Andreottola G
- Subjects
- Aerobiosis, Anaerobiosis, Bacteria enzymology, Biological Oxygen Demand Analysis, Nitrogen metabolism, Oxidation-Reduction, Sewage chemistry, Single-Cell Analysis, Bacteria growth & development, Bioreactors microbiology, Sewage microbiology, Waste Disposal, Fluid methods
- Abstract
In the Anaerobic Side-Stream Reactor (ASSR), part of the return sludge undergoes alternating aerobic and anaerobic conditions with the aim of reducing sludge production. In this paper, viability, enzymatic activity, death and lysis of bacterial cells exposed to aerobic and anaerobic conditions for 16 d were investigated at single-cell level by flow cytometry, with the objective of contributing to the understanding of the mechanisms of sludge reduction in the ASSR systems. Results indicated that total and viable bacteria did not decrease during the anaerobic phase, indicating that anaerobiosis at ambient temperature does not produce a significant cell lysis. Bacteria decay and lysis occurred principally under aerobic conditions. The aerobic decay rate of total bacteria (bTB) was considered as the rate of generation of lysed bacteria. Values of bTB of 0.07-0.11 d(-1) were measured in anaerobic + aerobic sequence. The enzymatic activity was not particularly affected by the transition from anaerobiosis to aerobiosis. Large solubilisation of COD and NH4(+) was observed only under anaerobic conditions, as a consequence of hydrolysis of organic matter, but not due to cell lysis. The observations supported the proposal of two independent mechanisms contributing equally to sludge reduction: (1) under anaerobic conditions: sludge hydrolysis of non-bacterial material, (2) under aerobic conditions: bacterial cell lysis and oxidation of released biodegradable compounds., (Copyright © 2015 Elsevier Ltd. All rights reserved.)
- Published
- 2015
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67. Micronucleus induction in mussels exposed to okadaic acid.
- Author
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Carvalho Pinto-Silva CR, Ferreira JF, Costa RH, Belli Filho P, Creppy EE, and Matias WG
- Subjects
- Animals, Environmental Monitoring methods, Micronucleus Tests, Time Factors, Bivalvia drug effects, Bivalvia genetics, Hemocytes drug effects, Marine Toxins toxicity, Okadaic Acid toxicity
- Abstract
Some toxins present in the marine environment are capable of inducing mutagenicity and/or carcinogenicity. Among these toxins, okadaic acid (OA) is gaining considerable interest since it induces DNA based modifications at low concentrations and accumulates in filter-feeding marine animals, including those used for human consumption. This study aims to evaluate the genotoxicity of OA in the haemocytes of the mussel Perna perna, using the micronucleus assay. Fifty-four mussels were separated into three groups of 18 animals. One group received 0.3 microg of OA diluted in 10 microl of ethanol and ultrapure water while the other groups were considered as controls and were exposed to a solvent plus seawater mixture. A significantly higher frequency of micronuclei was observed in haemocytes from the OA-exposed group. There were no statistical differences between the two control groups.
- Published
- 2003
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68. Human peroxisome proliferator-activated receptor alpha (PPARalpha) supports the induction of peroxisome proliferation in PPARalpha-deficient mouse liver.
- Author
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Yu S, Cao WQ, Kashireddy P, Meyer K, Jia Y, Hughes DE, Tan Y, Feng J, Yeldandi AV, Rao MS, Costa RH, Gonzalez FJ, and Reddy JK
- Subjects
- Animals, Cell Division drug effects, Gene Expression Regulation, Humans, Mice, Mice, Inbred C57BL, Mice, Knockout, Oxidation-Reduction, Peroxisomes drug effects, Pyrimidines pharmacology, RNA, Messenger analysis, Liver metabolism, Peroxisomes metabolism, Receptors, Cytoplasmic and Nuclear physiology, Transcription Factors physiology
- Abstract
Peroxisome proliferators, which function as peroxisome proliferator-activated receptor alpha (PPARalpha) agonists, induce peroxisomal, microsomal, and mitochondrial fatty acid oxidation enzymes, in conjunction with peroxisome proliferation, in liver cells. Sustained activation of PPARalpha leads to the development of liver tumors in rats and mice. The assertion that synthetic PPARalpha ligands pose negligible carcinogenic risk to humans is attributable, in part, to the failure to observe peroxisome proliferation in human hepatocytes. To explore the mechanism(s) of species-specific differences in response to PPARalpha ligands, we determined the functional competency of human PPARalpha in vivo and compared its potency with that of mouse PPARalpha. Recombinant adenovirus that expresses human or mouse PPARalpha was produced and administered intravenously to PPARalpha-deficient mice. Human as well as mouse PPARalpha fully restored the development of peroxisome proliferator-induced immediate pleiotropic responses, including peroxisome proliferation and enhanced expression of genes involved in lipid metabolism as well as nonperoxisomal genes, such as CD36, Ly-6D, Rbp7, monoglyceride lipase, pyruvate dehydrogenase kinase-4, and C3f, that have been identified recently to be up-regulated in livers with peroxisome proliferation. These studies establish that human PPARalpha is functionally competent and is equally as dose-sensitive as mouse PPARalpha in inducing peroxisome proliferation within the context of mouse liver environment and that it can heterodimerize with mouse retinoid X receptor, and this human PPARalpha-mouse retinoid X receptor chimeric heterodimer transcriptionally activates mouse PPARalpha target genes in a manner qualitatively similar to that of mouse PPARalpha.
- Published
- 2001
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69. DDB2 induces nuclear accumulation of the hepatitis B virus X protein independently of binding to DDB1.
- Author
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Nag A, Datta A, Yoo K, Bhattacharyya D, Chakrabortty A, Wang X, Slagle BL, Costa RH, and Raychaudhuri P
- Subjects
- G1 Phase, Hepatocytes metabolism, Humans, Protein Subunits, Tumor Cells, Cultured, Viral Regulatory and Accessory Proteins, Cell Nucleus metabolism, DNA-Binding Proteins metabolism, DNA-Binding Proteins physiology, Trans-Activators metabolism
- Abstract
The hepatitis B virus (HBV) X protein (HBx) is critical for the life cycle of the virus. HBx associates with several host cell proteins including the DDB1 subunit of the damaged-DNA binding protein DDB. Recent studies on the X protein encoded by the woodchuck hepadnavirus have provided correlative evidence indicating that the interaction with DDB1 is important for establishment of infection by the virus. In addition, the interaction with DDB1 has been implicated in the nuclear localization of HBx. Because the DDB2 subunit of DDB is required for the nuclear accumulation of DDB1, we investigated the role of DDB2 in the nuclear accumulation of HBx. Here we show that expression of DDB2 increases the nuclear levels of HBx. Several C-terminal deletion mutants of DDB2 that fail to bind DDB1 are able to associate with HBx, suggesting that DDB2 may associate with HBx independently of binding to DDB1. We also show that DDB2 enhances the nuclear accumulation of HBx independently of binding to DDB1, since a mutant that does not bind DDB1 is able to enhance the nuclear accumulation of HBx. HBV infection is associated with liver pathogenesis. We show that the nuclear levels of DDB1 and DDB2 are tightly regulated in hepatocytes. Studies with regenerating mouse liver indicate that during late G1 phase the nuclear levels of both subunits of DDB are transiently increased, followed by a sharp decrease in S phase. Taken together, these results suggest that DDB1 and DDB2 would participate in the nuclear functions of HBx effectively only during the late-G1 phase of the cell cycle.
- Published
- 2001
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70. Increased levels of forkhead box M1B transcription factor in transgenic mouse hepatocytes prevent age-related proliferation defects in regenerating liver.
- Author
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Wang X, Quail E, Hung NJ, Tan Y, Ye H, and Costa RH
- Subjects
- Aging genetics, Animals, Cell Division, Cyclin-Dependent Kinases metabolism, Cyclophilins genetics, DNA Replication, Forkhead Box Protein M1, Forkhead Transcription Factors, Hepatocytes physiology, Mice, Mice, Transgenic, Phosphorylation, RNA Probes, RNA, Antisense, Retinoblastoma Protein metabolism, Ribonucleases, S Phase, Transfection, Aging physiology, DNA-Binding Proteins genetics, Hepatocytes cytology, Liver physiology, Liver Regeneration physiology, Transcription Factors genetics
- Abstract
The forkhead box (Fox) family of transcription factors share homology in the winged helix/forkhead DNA-binding domain and play important roles in regulating cellular proliferation, differentiation, longevity, and cellular transformation. Forkhead box M1B (FoxM1B) is a ubiquitously expressed member of the Fox transcription factor family whose expression is restricted to proliferating cells and that mediates hepatocyte entry into DNA synthesis and mitosis during liver regeneration. Recent cDNA microarray studies indicated that age-related defects in cellular proliferation are associated with diminished expression of the FoxM1B transcription factor. Here, we show that increased levels of FoxM1B in regenerating liver of old transgenic mice restore the sharp peaks in hepatocyte DNA replication and mitosis that are the hallmarks of young regenerating mouse liver. Restoration of the young regenerating liver phenotype is associated with increased expression of numerous cell cycle regulatory genes that include cyclin D1, cyclin A2, cyclin F, cyclin B1, cyclin B2, Cdc25B, and p55cdc. Cotransfection assays in the human hepatoma HepG2 cell line demonstrated that FoxM1B protein stimulated expression of both the cyclin B1 and cyclin D1 promoters, suggesting that these cyclin genes are a direct FoxM1B transcriptional target. These results suggest that FoxM1B controls the transcriptional network of genes that are essential for cell division and exit from mitosis. Our results indicate that reduced expression of the FoxM1B transcription factor contributes to the decline in cellular proliferation observed in the aging process.
- Published
- 2001
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71. Defects in pulmonary vasculature and perinatal lung hemorrhage in mice heterozygous null for the Forkhead Box f1 transcription factor.
- Author
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Kalinichenko VV, Lim L, Stolz DB, Shin B, Rausa FM, Clark J, Whitsett JA, Watkins SC, and Costa RH
- Subjects
- Alleles, Animals, Apoptosis, Blotting, Western, Bone Morphogenetic Protein 4, Bone Morphogenetic Proteins metabolism, DNA, Complementary metabolism, Dose-Response Relationship, Drug, Endothelial Growth Factors metabolism, Endothelium metabolism, Forkhead Transcription Factors, Hemorrhage, Heterozygote, Immunohistochemistry, In Situ Nick-End Labeling, Lung pathology, Lymphokines metabolism, Mice, Mice, Knockout, Microscopy, Electron, Models, Genetic, Mutation, Oligonucleotide Array Sequence Analysis, Phenotype, Platelet Endothelial Cell Adhesion Molecule-1 metabolism, RNA, Messenger metabolism, Receptor Protein-Tyrosine Kinases metabolism, Receptors, Growth Factor metabolism, Receptors, Vascular Endothelial Growth Factor, Trans-Activators metabolism, Transcription Factors physiology, Vascular Endothelial Growth Factor A, Vascular Endothelial Growth Factors, beta-Galactosidase metabolism, DNA-Binding Proteins, Lung embryology, Lung metabolism, Transcription Factors biosynthesis, Transcription Factors genetics, Transcription Factors metabolism
- Abstract
Decreased pulmonary expression of Forkhead Box f1 (Foxf1) transcription factor was associated with lethal alveolar hemorrhage in 55% of the Foxf1 +/- newborn mice. The severity of the pulmonary abnormalities correlates with the levels of Foxf1 mRNA. Defects in alveolarization and vasculogenesis were observed in subsets of the Foxf1 +/- mice with relatively low levels of expression from the normal Foxf1 allele. Lung hemorrhage was coincident with disruption of the mesenchymal-epithelial cell interfaces in the alveolar and bronchiolar regions of the lung parenchyma and was associated with increased apoptosis and reduced surfactant protein B (SP-B) expression. Finally, the lung defect associated with the Foxf1 +/- mutation was accompanied by reduced expression of vascular endothelial growth factor (VEGF), the VEGF receptor 2 (Flk-1), bone morphogenetic protein 4 (Bmp-4), and the transcription factors of the Brachyury T-Box family (Tbx2-Tbx5) and Lung Kruppel-like Factor. Reduction in the level of Foxf1 caused neonatal pulmonary hemorrhage and abnormalities in alveologenesis, implicating this transcription factor in the regulation of mesenchyme-epithelial interaction critical for lung morphogenesis., (Copyright 2001 Academic Press.)
- Published
- 2001
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72. Earlier expression of the transcription factor HFH-11B diminishes induction of p21(CIP1/WAF1) levels and accelerates mouse hepatocyte entry into S-phase following carbon tetrachloride liver injury.
- Author
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Wang X, Hung NJ, and Costa RH
- Subjects
- Animals, Carbon Tetrachloride, Cell Adhesion Molecules metabolism, Cell Cycle Proteins metabolism, Chemical and Drug Induced Liver Injury, Cyclin-Dependent Kinase Inhibitor p21, Cyclins metabolism, Forkhead Box Protein M1, Forkhead Transcription Factors, Liver Diseases physiopathology, Male, Mice, Mice, Inbred Strains, Mice, Transgenic genetics, Time Factors, Transcription Factors metabolism, cdc25 Phosphatases metabolism, Cyclins genetics, Gene Expression physiology, Gene Expression Regulation physiology, Hepatocytes physiology, Liver Diseases genetics, Liver Diseases pathology, S Phase, Transcription Factors genetics
- Abstract
Partial hepatectomy (PH) or toxic liver injury induces the proliferation of terminally differentiated hepatic cells to regenerate the original size of the adult liver. Previous PH liver regeneration studies showed that premature transgenic expression of the Forkhead Box M1b (FoxM1b, HFH-11B) transcription factor accelerated hepatocyte entry into DNA replication (S-phase). In this study, we used carbon tetrachloride (CCl(4)) liver injury to induce a different type of mouse liver regeneration and show that premature hepatic HFH-11B levels also accelerate the onset of hepatocyte S-phase in this injury model. Unlike PH liver regeneration, earlier hepatocyte proliferation after CCl(4) liver injury is correlated with diminished transgenic hepatic levels of p21(CIP1/WAF1) at the G1/S transition of the cell cycle. Differential hybridization of cDNA arrays and RNase protection studies determined that CCl(4) regenerating liver of transgenic mice displayed early stimulated expression of the S-phase promoting cyclin D1 and cyclin E and sustained levels of Cdc25a phosphatase genes. Compared with previous PH liver regeneration studies, our data suggest that premature expression of HFH-11B activates distinct S-phase promotion pathways in the CCl(4) liver injury model. Although proliferating transgenic hepatocytes induced by either PH or CCl(4) liver injury displayed early expression of identical M-phase cyclin genes (cyclin B1, B2, A2, and F), only CCl(4) regenerating transgenic liver exhibited earlier expression of the M-phase promoting Cdc25b. These studies suggest that CCl(4) injury of transgenic liver not only uses the same mechanisms as PH to mediate accelerated hepatocyte entry into mitosis, but also promotes M-phase entry by stimulating Cdc25b expression.
- Published
- 2001
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73. Transcription factors in mouse lung development and function.
- Author
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Costa RH, Kalinichenko VV, and Lim L
- Subjects
- Amino Acid Motifs, Animals, Cell Differentiation, Epithelial Cells cytology, Epithelial Cells metabolism, Gene Expression Regulation, Developmental, Lung cytology, Mesoderm cytology, Mesoderm metabolism, Mice, Mice, Knockout, Mice, Transgenic, Morphogenesis physiology, Phenotype, Transcription Factors genetics, Lung embryology, Lung metabolism, Transcription Factors metabolism
- Abstract
Development of the mouse lung initiates on day 9.5 postcoitum from the laryngotracheal groove and involves mesenchymal-epithelial interactions, in particular, those between the splanchnic mesoderm and epithelial cells (derived from foregut endoderm) that induce cellular proliferation, migration, and differentiation, resulting in branching morphogenesis. This developmental process mediates formation of the pulmonary bronchiole tree and integrates a terminal alveolar region with an extensive endothelial capillary bed, which facilitates efficient gas exchange with the circulatory system. The major function of the mesenchymal-epithelial signaling is to potentiate the activity or expression of cell type-specific transcription factors in the developing lung, which, in turn, cooperatively bind to distinct promoter regions and activate target gene expression. In this review, we focus on the role of transcription factors in lung morphogenesis and the maintenance of differentiated gene expression. These lung transcription factors include forkhead box A2 [also known as hepatocyte nuclear factor (HNF)-3beta], HNF-3/forkhead homolog (HFH)-8 [also known as FoxF1 or forkhead-related activator-1], HNF-3/forkhead homolog-4 (also known as FoxJ1), thyroid transcription factor-1 (Nkx2.1), and homeodomain box A5 transcription factors, the zinc finger Gli (mouse homologs of the Drosophila cubitus interruptus) and GATA transcription factors, and the basic helix-loop-helix Pod1 transcription factor. We summarize the phenotypes of transgenic and knockout mouse models, which define important functions of these transcription factors in cellular differentiation and lung branching morphogenesis.
- Published
- 2001
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74. Differential expression of forkhead box transcription factors following butylated hydroxytoluene lung injury.
- Author
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Kalinichenko VV, Lim L, Shin B, and Costa RH
- Subjects
- Animals, Bronchi metabolism, Bronchi pathology, Butylated Hydroxytoluene, Cell Division physiology, Endothelium metabolism, Endothelium pathology, Forkhead Box Protein M1, Forkhead Transcription Factors, Hepatocyte Nuclear Factor 3-beta, Lung metabolism, Lung pathology, Lung Diseases chemically induced, Lung Diseases pathology, Male, Mesoderm pathology, Mice, Mice, Inbred BALB C, Muscle, Smooth cytology, Muscle, Smooth metabolism, DNA-Binding Proteins metabolism, Lung Diseases metabolism, Nuclear Proteins metabolism, Trans-Activators metabolism, Transcription Factors metabolism
- Abstract
The forkhead box (Fox) proteins are a growing family of transcription factors that have important roles in cellular proliferation and differentiation and in organ morphogenesis. The Fox family members hepatocyte nuclear factor (HNF)-3beta (Foxa2) and HNF-3/forkhead homolog (HFH)-8 (FREAC-1, Foxf1) are expressed in adult pulmonary epithelial and mesenchymal cells, respectively, but these cells display only low expression levels of the proliferation-specific HFH-11B gene (Trident, Foxm1b). The regulation of these Fox transcription factors in response to acute lung injury, however, has yet to be determined. We report here on the use of butylated hydroxytoluene (BHT)-mediated lung injury to demonstrate that HFH-11 protein and RNA levels were markedly increased throughout the period of lung repair. The maximum levels of HFH-11 were observed by day 2 following BHT injury when both bronchiolar and alveolar epithelial cells were undergoing extensive proliferation. Although BHT lung injury did not alter epithelial cell expression of HNF-3beta, a 65% reduction in HFH-8 mRNA levels was observed during the period of mesenchymal cell proliferation. HFH-8-expressing cells were colocalized with platelet endothelial cell adhesion molecule-1-positive alveolar endothelial cells and with alpha-smooth muscle actin-positive peribronchiolar smooth muscle cells.
- Published
- 2001
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75. Atypical mouse cerebellar development is caused by ectopic expression of the forkhead box transcription factor HNF-3beta.
- Author
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Zhou H, Hughes DE, Major ML, Yoo K, Pesold C, and Costa RH
- Subjects
- Animals, Animals, Newborn, Antigens, CD genetics, Apoptosis, Astrocytes cytology, Astrocytes metabolism, Basic Helix-Loop-Helix Transcription Factors, Cell Adhesion Molecules, Neuronal genetics, Cell Differentiation, Cell Movement, Cerebellum abnormalities, Cerebellum cytology, DNA-Binding Proteins genetics, DNA-Binding Proteins immunology, ErbB Receptors genetics, Extracellular Matrix Proteins genetics, Gene Expression, Genetic Markers genetics, Hepatocyte Nuclear Factor 3-beta, In Situ Hybridization, Insulin-Like Growth Factor Binding Protein 1 genetics, Insulin-Like Growth Factor Binding Protein 1 metabolism, Integrin alpha5, Mice, Mice, Transgenic, Nerve Tissue Proteins, Netrin Receptors, Neuroglia cytology, Neuroglia metabolism, Neurons cytology, Neurons metabolism, Nuclear Proteins genetics, Nuclear Proteins immunology, Oligonucleotide Array Sequence Analysis, Phenotype, Purkinje Cells cytology, Purkinje Cells metabolism, RNA, Messenger genetics, RNA, Messenger metabolism, Receptor, ErbB-4, Receptors, Nerve Growth Factor genetics, Reelin Protein, Serine Endopeptidases, Transcription Factors genetics, Cerebellum growth & development, Cerebellum metabolism, DNA-Binding Proteins metabolism, Nuclear Proteins metabolism
- Abstract
To assess the role of hepatocyte nuclear factor-3beta (HNF-3beta) in hepatocyte-specific gene transcription, we reported the characterization of the liver phenotype with transgenic mice in which the -3-kb transthyretin (TTR) promoter functioned to increase HNF-3beta expression. During breeding of the TTR-HNF-3beta transgenic mice we noticed that they displayed severe ataxia. In this study, we describe the analysis of our transgenic cerebellar phenotype and demonstrate that ectopic expression of HNF-3beta disrupted cerebellar morphogenesis and caused reduction in cerebellar size. In postnatal cerebellum, the HNF-3beta transgene expression pattern is colocalized to glial fibrillary acidic protein-positive cerebellar astrocytes and Bergmann glial cells. As a result of protracted expression, the transgenic cerebella are impaired in terms of astrocyte dispersal and formation of Bergmann glial cell processes. This caused a disruption in neuronal cell migration to the cortical laminar layers and Purkinje dendritic arbor maturation, thus leading to diminished foliation. Differential hybridization of cDNA arrays was used to identify altered expression of cerebellar genes, which is consistent with the observed defect in transgenic cerebellar morphogenesis and size as well as glial maturation. These include diminished expression of the brain lipid-binding protein, which is required for glial morphological differentiation, and the basic helix-loop-helix NeuroD/Beta2 and homeodomain Engrailed-2 transcription factors, which are required for normal cerebellar morphogenesis and foliation. Undetectable levels of ataxia telangiectasia (ATM), which is required for proper development of the Purkinje dendritic arbor, were found in postnatal transgenic cerebella. Furthermore, the transgenic cerebella displayed levels of insulin-like growth factor binding protein-1 elevated to 22 times greater than those measured for wild-type cerebella, an elevation consistent with the reduction in transgenic cerebellar size.
- Published
- 2001
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76. Adenovirus-mediated increase of HNF-3 levels stimulates expression of transthyretin and sonic hedgehog, which is associated with F9 cell differentiation toward the visceral endoderm lineage.
- Author
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Tan Y, Costa RH, Kovesdi I, and Reichel RR
- Subjects
- Animals, Cell Line, Cell Lineage, Chloramphenicol O-Acetyltransferase genetics, DNA-Binding Proteins metabolism, Endoderm, Genetic Vectors, Hedgehog Proteins, Hepatocyte Nuclear Factor 3-alpha, Hepatocyte Nuclear Factor 3-beta, Humans, Mice, Nuclear Proteins metabolism, RNA, Messenger genetics, RNA, Messenger metabolism, Tumor Cells, Cultured, Adenoviridae genetics, Cell Differentiation genetics, DNA-Binding Proteins biosynthesis, Gene Expression Regulation genetics, Nuclear Proteins biosynthesis, Prealbumin genetics, Trans-Activators genetics, Transcription Factors
- Abstract
Retinoic acid-induced differentiation of mouse F9 embryonal carcinoma cells toward the visceral endoderm lineage is accompanied by increased expression of the Forkhead Box (Fox) transcription factors hepatocyte nuclear factor 3a (HNF-3alpha) and HNF-3beta, suggesting that they play a crucial role in visceral endoderm development. Retinoic acid stimulation results in a cascade of HNF-3 induction in which HNF-3alpha is a primary target for retinoic acid action and its increase is required for subsequent induction of HNF-3beta expression. Increased expression of HNF-3beta precedes activation of its known target genes, including transthyretin (TTR), Sonic hedgehog (Shh), HNF-1alpha, HNF-1beta, and HNF-4alpha. In order to examine whether increased HNF-3 expression is sufficient to induce expression of its downstream target genes without retinoic acid stimulation, we have used adenovirus-based expression vectors to increase HNF-3 protein levels in F9 cells. We demonstrate that adenovirus-mediated increase of HNF-3alpha levels in F9 cells is sufficient to induce activation of endogenous HNF-3beta levels followed by increased TTR and Shh expression. Furthermore, we show that elevated HNF-3beta levels stimulate expression of endogenous TTR and Shh without retinoic acid stimulation. Moreover, ectopic HNF-3 levels in undifferentiated F9 cells are insufficient to induce HNF-3alpha, HNF-1alpha, HNF-1beta, and HNF-4alpha expression, suggesting that their transcriptional activation required other regulatory proteins induced by the retinoic acid differentiation program. Finally, our studies demonstrate the utility of cell infections with adenovirus expressing distinct transcription factors to identify endogenous target genes, which are assembled with the appropriate nucleosome structure.
- Published
- 2001
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77. Odor control of an anaerobic lagoon with a biological cover: floating peat beds.
- Author
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Picot B, Paing J, Toffoletto L, Sambuco JP, and Costa RH
- Subjects
- Air Pollution prevention & control, Bacteria, Anaerobic physiology, Biodegradation, Environmental, Oxidation-Reduction, Plants, Sulfur chemistry, Waste Disposal, Fluid, Hydrogen Sulfide analysis, Odorants, Soil
- Abstract
The use of a biological cover for in situ control of gaseous sulfide emission from an anaerobic pond was investigated by a laboratory-scale experiment. The biological cover, constituting by a peat bed floating on the wastewater, caused a reduction of the H2S emission rate by 84.6%. The addition of Fe3+ (with FeCl3) and plants (Juncus effusus L.) to the peat bed significantly improved the performance to reach a H2S removal of 95.5%. Despite the fluctuations in the sulfide concentration in the wastewater, the performance of the biological covers remained constant during the entire period of the study. The analysis of the different forms of sulfur accumulated in the peat beds allowed the understanding of the mechanisms involved in H2S removal. The high amount of sulfate demonstrated that the conditions were favorable to the biological oxidation of H2S. The addition of Fe3+ increased the formation of insoluble ferrous monosulfide (FeS) and pyrite (FeS2). The plants seemed to convert sulfate into elemental and organic sulfur.
- Published
- 2001
78. Elevated levels of hepatocyte nuclear factor 3beta in mouse hepatocytes influence expression of genes involved in bile acid and glucose homeostasis.
- Author
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Rausa FM, Tan Y, Zhou H, Yoo KW, Stolz DB, Watkins SC, Franks RR, Unterman TG, and Costa RH
- Subjects
- ATP Binding Cassette Transporter, Subfamily B metabolism, ATP-Binding Cassette Transporters metabolism, Animals, Base Sequence, Bile Acids and Salts metabolism, Blotting, Western, Carrier Proteins metabolism, Cell Line, DNA Methylation, Glucose metabolism, Glutathione Transferase metabolism, Glycogen metabolism, Hepatocyte Nuclear Factor 3-beta, Hepatocyte Nuclear Factor 6, Homeodomain Proteins metabolism, Immunohistochemistry, Insulin-Like Growth Factor Binding Protein 1 blood, Insulin-Like Growth Factor Binding Protein 1 metabolism, Ligands, Liver embryology, Liver metabolism, Liver pathology, Mice, Mice, Transgenic, Microscopy, Electron, Models, Genetic, Molecular Sequence Data, Organic Anion Transporters, Sodium-Dependent, Phenotype, Prealbumin genetics, Prealbumin metabolism, Promoter Regions, Genetic, Protein Isoforms, Recombinant Proteins metabolism, Symporters, Time Factors, Trans-Activators metabolism, Transcription, Genetic, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, Liver cytology, Membrane Transport Proteins, Nuclear Proteins genetics, Nuclear Proteins metabolism, Transcription Factors
- Abstract
The winged helix transcription factor, hepatocyte nuclear factor-3beta (HNF-3beta), mediates the hepatocyte-specific transcription of numerous genes important for liver function. However, the in vivo role of HNF-3beta in regulating these genes remains unknown because homozygous null HNF3beta mouse embryos die in utero prior to liver formation. In order to examine the regulatory function of HNF-3beta, we created transgenic mice in which the -3-kb transthyretin promoter functions to increase hepatocyte expression of the rat HNF-3beta protein. Postnatal transgenic mice exhibit growth retardation, depletion of hepatocyte glycogen storage, and elevated levels of bile acids in serum. The retarded growth phenotype is likely due to a 20-fold increase in hepatic expression of insulin-like growth factor binding protein 1 (IGFBP-1), which results in elevated levels in serum of IGFBP-1 and limits the biological availability of IGFs required for postnatal growth. The defects in glycogen storage and serum bile acids coincide with diminished postnatal expression of hepatocyte genes involved in gluconeogenesis (phosphoenolpyruvate carboxykinase and glycogen synthase) and sinusoidal bile acid uptake (Ntcp), respectively. These changes in gene transcription may result from the disruptive effect of HNF-3beta on the hepatic expression of the endogenous mouse HNF-3alpha,-3beta, -3gamma, and -6 transcription factors. Furthermore, adult transgenic livers lack expression of the canalicular phospholipid transporter, mdr2, which is consistent with ultrastructure evidence of damage to transgenic hepatocytes and bile canaliculi. These transgenic studies represent the first in vivo demonstration that the HNF-3beta transcriptional network regulates expression of hepatocyte-specific genes required for bile acid and glucose homeostasis, as well as postnatal growth.
- Published
- 2000
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79. Angiotensin converting-like enzymes from urine of untreated renovascular hypertensive and normal patients: purification and characterization.
- Author
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Costa RH, Casarini DE, Plavnik FL, Marson O, and Alves KB
- Subjects
- Adult, Angiotensin-Converting Enzyme Inhibitors pharmacology, Female, Humans, Kinetics, Male, Molecular Weight, Peptidyl-Dipeptidase A isolation & purification, Hypertension, Renovascular enzymology, Peptidyl-Dipeptidase A urine
- Abstract
Angiotensin converting-like enzymes (ACE) were isolated from urine of normal (P0N, P1N and P2N) and untreated renovascular hypertensive (P0, P1 and P2) patients. The urine were submitted to ion exchange chromatography. Enzymes P0 and P0N were eluted with the equilibrium buffer (0.02 M Tris-HCl, pH 7.0), while P1, P1N, P2 and P2N with ionic strength linear gradient of 0.02-0.5 M Tris-HCl, pH 7.0 in 0.7 mS and P2 and P2N in 1.2 mS conductance. The active fractions were submitted to gel filtration in Sephadex G-150, equilibrated and performed with 0.05 M Tris-HCl/0.15 M NaCl buffer, pH 8.0. All enzymes were homogeneous when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) (molecular mass: P0, P2 and P2N about 60 kDa; P1, 95 kDa and P21N 170 kDa). The enzymes were recognized by Y1 polyclonal antibody raised against human renal ACE. The K(M) values were in millimolar order for hippuryl-L-His-Leu (HHL) while for benzyloxycarbonyl-Phe-L-His-Leu (ZFHL) they were in 10(-4) M order. The enzymes were able to hydrolyze angiotensin I (AI) (P0 and P0N about 25%, P1 and P1N about 70%, P2 100% and P2N 66%) and bradykinin (BK) (P0N 22%, P1N 81%, P2N 62%, P0 and P1 50% and P2 35%), and their activities were inhibited by captopril.
- Published
- 2000
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80. The nuclear receptor fetoprotein transcription factor is coexpressed with its target gene HNF-3beta in the developing murine liver, intestine and pancreas.
- Author
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Rausa FM, Galarneau L, Bélanger L, and Costa RH
- Subjects
- Animals, Carcinoma, Hepatocellular genetics, DNA-Binding Proteins metabolism, Endoderm, Gene Expression Regulation, Developmental, Hepatocyte Nuclear Factor 3-beta, Intestines growth & development, Liver growth & development, Mice, Nuclear Proteins metabolism, Pancreas growth & development, Receptors, Cytoplasmic and Nuclear genetics, Receptors, Cytoplasmic and Nuclear metabolism, Transcription Factors metabolism, Tumor Cells, Cultured, DNA-Binding Proteins genetics, Intestines embryology, Liver embryology, Nuclear Proteins genetics, Pancreas embryology, Transcription Factors genetics
- Abstract
During organogenesis, the winged helix hepatocyte nuclear factor 3beta (HNF-3beta) protein participates in regulating gene transcription in the developing esophagus, trachea, liver, lung, pancreas, and intestine. Hepatoma cell transfection studies identified a critical HNF-3beta promoter factor, named UF2-H3beta, and here, we demonstrate that UF2-H3beta is identical to the fetoprotein transcription factor (FTF). In situ hybridization studies of mouse embryos demonstrate that FTF expression initiates in the foregut endoderm during liver and pancreatic morphogenesis (day 9) and that earlier expression of FTF is observed in the yolk sac endoderm, branchial arch and neural crest cells (day 8). Abundant FTF hybridization signals are observed throughout morphogenesis of the liver, pancreas, and intestine and its expression continues in the epithelial cells of these adult organs. In day 17 mouse embryos and adult pancreas, however, expression of FTF becomes restricted to the exocrine acinar and ductal epithelial cells.
- Published
- 1999
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81. Premature expression of the winged helix transcription factor HFH-11B in regenerating mouse liver accelerates hepatocyte entry into S phase.
- Author
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Ye H, Holterman AX, Yoo KW, Franks RR, and Costa RH
- Subjects
- Animals, CCAAT-Enhancer-Binding Proteins, Cell Nucleus metabolism, Cyclins biosynthesis, Cyclins genetics, DNA Replication, DNA-Binding Proteins biosynthesis, DNA-Binding Proteins genetics, Forkhead Box Protein M1, Forkhead Transcription Factors, Humans, Liver cytology, Male, Mice, Mice, Transgenic, Mitosis, Nuclear Proteins biosynthesis, Phosphoproteins biosynthesis, Phosphoproteins genetics, Recombinant Fusion Proteins biosynthesis, Recombinant Fusion Proteins genetics, S Phase, Signal Transduction, Time Factors, Transcription Factors genetics, X-ray Repair Cross Complementing Protein 1, Liver metabolism, Liver Regeneration physiology, Transcription Factors biosynthesis
- Abstract
Two-thirds partial hepatectomy (PH) induces differentiated cells in the liver remnant to proliferate and regenerate to its original size. The proliferation-specific HNF-3/fork head homolog-11B protein (HFH-11B; also known as Trident and Win) is a family member of the winged helix/fork head transcription factors and in regenerating liver its expression is reactivated prior to hepatocyte entry into DNA replication (S phase). To examine whether HFH-11B regulates hepatocyte proliferation during liver regeneration, we used the -3-kb transthyretin (TTR) promoter to create transgenic mice that displayed ectopic hepatocyte expression of HFH-11B. Liver regeneration studies with the TTR-HFH-11B mice demonstrate that its premature expression resulted in an 8-h acceleration in the onset of hepatocyte DNA replication and mitosis. This liver regeneration phenotype is associated with protracted expression of cyclin D1 and C/EBPbeta, which are involved in stimulating DNA replication and premature expression of M phase promoting cyclin B1 and cdc2. Consistent with the early hepatocyte entry into S phase, regenerating transgenic livers exhibited earlier expression of DNA repair genes (XRCC1, mHR21spA, and mHR23B). Furthermore, in nonregenerating transgenic livers, ectopic HFH-11B expression did not elicit abnormal hepatocyte proliferation, a finding consistent with the retention of the HFH-11B transgene protein in the cytoplasm. We found that nuclear translocation of the HFH-11B transgene protein requires mitogenic signalling induced by PH and that its premature availability in regenerating transgenic liver allowed nuclear translocation to occur 8 h earlier than in wild type.
- Published
- 1999
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82. HNF-3/forkhead homologue-4 influences lung morphogenesis and respiratory epithelial cell differentiation in vivo.
- Author
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Tichelaar JW, Lim L, Costa RH, and Whitsett JA
- Subjects
- Animals, Basic Helix-Loop-Helix Leucine Zipper Transcription Factors, Cell Differentiation physiology, Cells, Cultured, Epithelial Cells cytology, Epithelial Cells physiology, Gene Expression Regulation, Developmental physiology, Hepatocyte Nuclear Factor 4, Lung cytology, Mice, Mice, Transgenic, Morphogenesis physiology, Respiratory System cytology, DNA-Binding Proteins, Lung embryology, Phosphoproteins physiology, Transcription Factors physiology
- Abstract
HNF-3/forkhead homologue 4 (HFH-4), a transcription factor of the winged helix/forkhead family, is expressed in various tissues including lung, brain, oviduct, testis, and embryonic kidney. In order to test whether the temporospatial expression of HFH-4 influences lung morphogenesis, HFH-4 was expressed in lungs of transgenic mice under control of the surfactant protein C (SP-C) promoter. The morphology of the lungs from SP-C/HFH-4 embryos (day 18 postconception) was distinctly abnormal, and the severity of the alterations correlated with the level of transgene expression as detected by in situ hybridization. At high levels of expression, HFH-4 altered epithelial cell differentiation and inhibited branching morphogenesis. Atypical cuboidal or columnar cells lined the lung periphery of SP-C/HFH-4 transgenic mice. The atypical epithelial cells seen in the SP-C/HFH-4 mice expressed thyroid transcription factor-1 and hepatocyte nuclear factor 3beta (HNF-3beta). However, surfactant proteins SP-B, SP-C, and Clara cell secretory protein, normally produced by nonciliated epithelial cells in lung parenchyma were lacking. beta-Tubulin IV, a marker of ciliated cells, stained the atypical columnar cells produced by expression of high levels of the SP-C/HFH-4 transgene. Ectopic expression of HFH-4 in developing mouse lung altered epithelial cell differentiation and morphology, restricting the expression of markers typical of nonciliated cells of the distal lung parenchyma., (Copyright 1999 Academic Press.)
- Published
- 1999
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83. Genesis, a Winged Helix transcriptional repressor, has embryonic expression limited to the neural crest, and stimulates proliferation in vitro in a neural development model.
- Author
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Hromas R, Ye H, Spinella M, Dmitrovsky E, Xu D, and Costa RH
- Subjects
- Animals, Cell Differentiation genetics, Cell Division genetics, Embryonic and Fetal Development genetics, Forkhead Transcription Factors, Helix-Turn-Helix Motifs genetics, Mice, Organ Specificity, Trans-Activators genetics, DNA-Binding Proteins genetics, Gene Expression Regulation, Developmental, Neural Crest embryology, Repressor Proteins genetics
- Abstract
A novel repressor of the Winged Helix (formerly HNF-3/Forkhead) transcriptional regulatory family, termed Genesis (also called HFH2), was previously found to be exclusively expressed in primitive embryonic cell lines. In this study in situ cRNA hybridization experiments revealed that Genesis was expressed during embryogenesis only in developing neural crest cells. Its expression diminished upon their terminal differentiation into sympathetic and parasympathetic neurons. Based on that finding, Genesis was retrovirally transduced into pluripotent N-Tera-2 clone D1 (NT2/D1) teratocarcinoma cells, which are a well-described in vitro model of neural development. Retinoic acid (RA) treatment will drive these cells to differentiation toward the neuronal lineage and cause an increase in expression of the cyclin-dependent kinase inhibitor p21 protein, which leads to an inhibition in cellular proliferation. Although RA-induced expression of neuronal differentiation markers was not influenced by forced overexpression of Genesis in NT2-D1 cells, proliferation of Genesis-transduced cells continued following RA treatment. RA was unable to induce the expression of the cyclin-dependent kinase inhibitor p21 in the Genesis-transduced cells, but Go/G1 tumor suppressor p53 expression was induced normally. Therefore, Genesis may play a role in the regulation of primitive neural crest development by preventing terminal quiescence through inhibition of p21 protein expression. These data also lend evidence for the hypothesis that proliferation and differentiation pathways are not irrevocably linked, but can function independently.
- Published
- 1999
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84. HNF-3/forkhead homologue-4 (HFH-4) is expressed in ciliated epithelial cells in the developing mouse lung.
- Author
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Tichelaar JW, Wert SE, Costa RH, Kimura S, and Whitsett JA
- Subjects
- Animals, Basic Helix-Loop-Helix Leucine Zipper Transcription Factors, DNA-Binding Proteins biosynthesis, DNA-Binding Proteins metabolism, Epithelial Cells metabolism, Hepatocyte Nuclear Factor 4, Immunohistochemistry, Mice, Mice, Knockout, Nuclear Proteins biosynthesis, Nuclear Proteins genetics, Phosphoproteins metabolism, Protein Biosynthesis, Thyroid Nuclear Factor 1, Tissue Distribution, Transcription Factors genetics, Transcription Factors metabolism, Tubulin biosynthesis, Tubulin metabolism, Lung embryology, Lung metabolism, Phosphoproteins biosynthesis, Transcription Factors biosynthesis, Uteroglobin
- Abstract
HNF-3/forkhead homologue-4 (HFH-4), a transcription factor of the winged-helix/forkhead family, was detected by immunohistochemistry in tissue of the developing mouse. HFH-4 protein was present in epithelial cells of the lung, trachea, oviduct, and embryonic esophagus, and in ependymal cells lining the spinal column and ventricles of the brain. In lung, trachea, and nose, HFH-4 was expressed in a distinct subset of epithelial cells that also expressed beta-tubulin IV, a ciliated cell marker. Cellular sites of HFH-4 and beta-tubulin IV expression were distinct from that of Clara cell secretory protein (CCSP), which was detected in nonciliated epithelial cells in the conducting airway of the lung. HFH-4 and beta-tubulin IV, but not CCSP, were detected in the respiratory epithelium of thyroid transcription factor-1 (TTF-1) gene-targeted mice. The presence of HFH-4 and beta-tubulin IV in TTF-1 gene-targeted mice demonstrates that differentiation of ciliated epithelium does not require TTF-1. Co-localization of HFH-4 and beta-tubulin IV staining in various tissues during mouse development supports a role for HFH-4 in the differentiation of ciliated cell lineages.
- Published
- 1999
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85. The -4 kilobase promoter region of the winged helix transcription factor HNF-3alpha gene elicits transgene expression in mouse embryonic hepatic and intestinal diverticula.
- Author
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Clevidence DE, Zhou H, Lau LF, and Costa RH
- Subjects
- Animals, Embryonic and Fetal Development genetics, Hepatocyte Nuclear Factor 3-alpha, Lung embryology, Mice, Mice, Transgenic, Notochord embryology, Organ Specificity, Recombinant Fusion Proteins, DNA-Binding Proteins genetics, Gene Expression Regulation, Developmental genetics, Intestines embryology, Nuclear Proteins genetics, Promoter Regions, Genetic genetics, Transcription Factors genetics
- Abstract
Murine hepatocyte nuclear factor-3alpha (HNF-3alpha) protein is a member of a large family of developmentally regulated transcription factors that share homology in the winged helix/fork head DNA binding domain and participate in embryonic pattern formation. HNF-3alpha also mediates cell-specific transcription of genes important for the function of hepatocytes, intestinal, pancreatic and bronchiolar epithelium. We have previously determined that -520 nucleotides upstream of the rat HNF-3alpha gene were sufficient to elicit hepatoma-specific expression in transfection assays and reported on a novel HNF-3alpha expression pattern in the renal pelvis urothelium of the embryonic and adult kidney. We also showed that retinoic acid mediated activation of the HNF-3alpha gene required -4 kb of the HNF-3alpha promoter region in F9 teratocarcinoma transfections. In order to determine regulatory sequences mediating the HNF-3alpha cellular expression pattern in developing mouse embryos, we created transgenic mice bearing the -4 kb HNF-3alpha promoter region driving expression of the beta-galactosidase transgene. Embryonic analysis of two transgenic mouse lines demonstrated that the -4 kb HNF-3alpha promoter sequences were sufficient to elicit transgene expression in the developing liver, intestine, esophagus, nasal epithelial cells and floorplate of the neurotube, but not in the mesodermal notochord or in the lung bud. One of the transgenic lines also exhibited proper expression in the mesonephric ducts and metanephric diverticulum, suggesting that the -4 kb HNF-3alpha promoter region contained a subset of the regulatory sequences necessary for HNF-3alpha expression in the developing kidney.
- Published
- 1998
86. In situ hybridization with 33P-labeled RNA probes for determination of cellular expression patterns of liver transcription factors in mouse embryos.
- Author
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Rausa FM, Ye H, Lim L, Duncan SA, and Costa RH
- Subjects
- Animals, Cell Differentiation physiology, DNA-Binding Proteins genetics, Gestational Age, Hepatocyte Nuclear Factor 3-beta, Hepatocyte Nuclear Factor 6, Homeodomain Proteins genetics, Intestines embryology, Liver embryology, Mice, Nuclear Proteins genetics, Oligonucleotides, Antisense genetics, Pancreas embryology, Phosphorus Isotopes, Prealbumin genetics, Promoter Regions, Genetic, RNA, Messenger analysis, Trans-Activators genetics, Gene Expression Regulation, Developmental genetics, In Situ Hybridization, Fluorescence methods, RNA Probes genetics, Transcription Factors analysis
- Abstract
Murine hepatocyte nuclear factor-3beta (HNF-3beta) protein is a member of a large family of developmentally regulated transcription factors that share homology in the winged helix/fork head DNA binding domain and that participate in embryonic pattern formation. HNF-3beta also mediates cell-specific transcription of genes important for the function of hepatocytes, intestinal and bronchiolar epithelium, and pancreatic acinar cells. We have previously identified a hepatocyte and pancreatic cut-homeodomain transcription factor, HNF-6, which is required for HNF-3beta promoter activity. In this study, we used in situ hybridization studies of stage-specific embryos to demonstrate that HNF-6 and its target gene, HNF-3beta, are coexpressed in the foregut endoderm and in the pancreatic and hepatic diverticulum. More detailed analysis of HNF-6 and HNF-3beta's developmental expression patterns provides evidence of colocalization in hepatocytes, intestinal epithelium, and pancreatic ductal epithelium and exocrine acinar cells. In support of the role of HNF-6 in regulating HNF-3beta expression in developing hepatocytes, their liver expression levels are both transiently reduced between 14 and 15 days of gestation. At day 18 of gestation and in adult pancreas, HNF-6 and HNF-3beta transcripts remain colocalized in the exocrine acinar cells, but their expression patterns diverge in endocrine cells. HNF-3beta expression is restricted to the endocrine cells of the islets of Langerhans, whereas the ductal epithelium expresses HNF-6. We discuss these expression patterns with respect to specification of hepatocytes and differentiation of the endocrine and exocrine pancreas., (Copyright 1998 Academic Press.)
- Published
- 1998
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87. The cut-homeodomain transcriptional activator HNF-6 is coexpressed with its target gene HNF-3 beta in the developing murine liver and pancreas.
- Author
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Rausa F, Samadani U, Ye H, Lim L, Fletcher CF, Jenkins NA, Copeland NG, and Costa RH
- Subjects
- Amino Acid Sequence, Animals, Carcinoma, Hepatocellular pathology, Cell Differentiation, Crosses, Genetic, DNA-Binding Proteins biosynthesis, Female, Fetal Proteins biosynthesis, Fetal Proteins genetics, Hepatocyte Nuclear Factor 3-beta, Hepatocyte Nuclear Factor 6, Homeodomain Proteins biosynthesis, Homeodomain Proteins genetics, Humans, In Situ Hybridization, Intestinal Mucosa embryology, Intestinal Mucosa metabolism, Islets of Langerhans embryology, Islets of Langerhans metabolism, Liver metabolism, Liver Neoplasms pathology, Male, Mice, Mice, Inbred C57BL, Molecular Sequence Data, Multigene Family, Muridae genetics, Nuclear Proteins biosynthesis, Organ Specificity, Pancreas metabolism, Promoter Regions, Genetic, Sequence Alignment, Sequence Homology, Amino Acid, Trans-Activators biosynthesis, Trans-Activators genetics, Transcription, Genetic, Tumor Cells, Cultured, DNA-Binding Proteins genetics, Fetal Proteins physiology, Gene Expression Regulation, Developmental, Genes, Homeobox, Homeodomain Proteins physiology, Liver embryology, Nuclear Proteins genetics, Pancreas enzymology, Trans-Activators physiology, Transcription Factors
- Abstract
Murine hepatocyte nuclear factor-3 beta (HNF-3 beta) protein is a member of a large family of developmentally regulated transcription factors that share homology in the winged helix/fork head DNA binding domain and that participate in embryonic pattern formation. HNF-3 beta also mediates cell-specific transcription of genes important for the function of hepatocytes, intestinal and bronchiolar epithelial, and pancreatic acinar cells. We have previously identified a liver-enriched transcription factor, HNF-6, which is required for HNF-3 beta promoter activity and also recognizes the regulatory region of numerous hepatocyte-specific genes. In this study we used the yeast one-hybrid system to isolate the HNF-6 cDNA, which encodes a cut-homeodomain-containing transcription factor that binds with the same specificity as the liver HNF-6 protein. Cotransfection assays demonstrate that HNF-6 activates expression of a reporter gene driven by the HNF-6 binding site from either the HNF-3 beta or transthyretin (TTR) promoter regions. We used interspecific backcross analysis to determine that murine Hnf6 gene is located in the middle of mouse chromosome 9. In situ hybridization studies of staged specific embryos demonstrate that HNF-6 and its potential target gene, HNF-3 beta, are coexpressed in the pancreatic and hepatic diverticulum. More detailed analysis of HNF-6 and HNF-3 beta's developmental expression patterns provides evidence of colocalization in hepatocytes, intestinal epithelial, and in the pancreatic ductal epithelial and exocrine acinar cells. The expression patterns of these two transcription factors do not overlap in other endoderm-derived tissues or the neurotube. We also found that HNF-6 is also abundantly expressed in the dorsal root ganglia, the marginal layer, and the midbrain. At day 18 of gestation and in the adult pancreas, HNF-6 and HNF-3 beta transcripts colocalize in the exocrine acinar cells, but their expression patterns diverge in other pancreatic epithelium. HNF-6, but not HNF-3 beta, expression continues in the pancreatic ductal epithelium, whereas only HNF-3 beta becomes restricted to the endocrine cells of the islets of Langerhans. We discuss these expression patterns with respect to specification of hepatocytes and differentiation of the endocrine and exocrine pancreas.
- Published
- 1997
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88. Hepatocyte nuclear factor-3beta limits cellular diversity in the developing respiratory epithelium and alters lung morphogenesis in vivo.
- Author
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Zhou L, Dey CR, Wert SE, Yan C, Costa RH, and Whitsett JA
- Subjects
- Animals, Cadherins genetics, Cell Differentiation, DNA-Binding Proteins genetics, Endothelial Growth Factors genetics, Hepatocyte Nuclear Factor 3-beta, Humans, Lung cytology, Lymphokines genetics, Mesoderm cytology, Mice, Mice, Transgenic, Morphogenesis, Nuclear Proteins genetics, RNA, Messenger metabolism, Rats, Transcription Factors genetics, Vascular Endothelial Growth Factor A, Vascular Endothelial Growth Factors, DNA-Binding Proteins physiology, Epithelial Cells cytology, Lung embryology, Nuclear Proteins physiology, Transcription Factors physiology
- Abstract
Hepatocyte nuclear factor-3beta (HNF-3beta), a nuclear protein of the winged helix family of transcription factors, is known to play a critical role in the formation of the embryonic node, notochord, and foregut endoderm. HNF-3beta influences the expression of a number of target genes in the respiratory epithelium, activating transcription of thyroid transcription factor-1, surfactant protein-B and clara cell secretory protein. In order to discern the role of HNF-3beta in differentiation and gene expression in the lung, HNF-3beta was expressed in developing respiratory epithelial cells of transgenic mice, under the control of the human surfactant protein C gene promoter. Pulmonary abnormalities were observed in the lungs of fetal mice bearing the HNF-3beta transgene. Differentiation of distal respiratory epithelial cells was arrested in the early pseudoglandular stage. Branching morphogenesis and vasculogenesis were markedly disrupted in association with decreased E-cadherin and vascular endothelial growth factor expression. HNF-3beta limits cellular diversity of developing respiratory epithelium and alters lung morphogenesis in vivo, suggesting that precise temporal-spatial regulation of HNF-3beta expression is critical for respiratory epithelial cell differentiation and lung morphogenesis.
- Published
- 1997
- Full Text
- View/download PDF
89. The winged helix transcription factor HFH-4 is expressed during choroid plexus epithelial development in the mouse embryo.
- Author
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Lim L, Zhou H, and Costa RH
- Subjects
- Amino Acid Sequence, Animals, Bronchi embryology, Bronchi metabolism, Choroid Plexus embryology, DNA-Binding Proteins genetics, Epithelium embryology, Epithelium metabolism, Humans, Mice, Molecular Sequence Data, Choroid Plexus metabolism, Gene Expression Regulation, Developmental, Transcription Factors genetics
- Abstract
Mammalian hepatocyte nuclear factor-3 (HNF-3) and the Drosophila homeotic gene fork head proteins are prototypes of an extensive family of cell-specific transcription factors that share homology in the winged helix DNA-binding domain. One of these mammalian family members, HNF-3/fork head homolog-4 (HFH-4), was isolated by PCR amplification of rodent brain cDNA and exhibits abundant expression in the adult bronchiolar epithelium. In this study, we performed in situ hybridization of stage-specific mouse embryos and report on a novel expression pattern of the HFH-4 gene in both the presumptive and differentiated choroid plexus epithelium, which is responsible for the synthesis and secretion of cerebrospinal fluid (CSF) proteins. We also showed that HFH-4 is a potent transcriptional activator in cotransfection assays and defined several protein sequences important for HFH-4 transcriptional activity. We used in vitro DNA-binding site selection with recombinant HFH-4 protein and determined that the HFH-4 protein recognizes the DNA consensus sequences HWDTGTTTGTTTA or KTTTGTTGTTKTW (where H is not G, W is A or T, D is not C, and K is G or T). We used this HFH-4 consensus to identify potential HFH-4 target genes in the choroid plexus epithelium and demonstrated that these promoter sequences bind to recombinant HFH-4 protein in electrophoretic mobility shift assays. Recombinant HFH-4 formed specific protein-DNA complexes with the promoter regions of the human prothrombin, beta amyloid precursor protein, alpha1-antichymotrypsin, cystic fibrosis transmembrane conductance regulator and rodent alpha2-macroglobulin, growth hormone receptors, and insulin-like growth factor II genes. Furthermore, we identified putative HFH-4 target genes in the bronchiolar epithelium including the clara cell secretory protein gene and the HNF-3alpha gene, a winged helix family member involved in the transcriptional regulation of genes in the bronchiolar epithelium. In support of these binding studies, cotransfection assays show that HFH-4 potentiates expression of the HNF-3alpha and clara cell secretory protein promoter regions.
- Published
- 1997
- Full Text
- View/download PDF
90. Hepatocyte nuclear factor-3 alpha promoter regulation involves recognition by cell-specific factors, thyroid transcription factor-1, and autoactivation.
- Author
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Peterson RS, Clevidence DE, Ye H, and Costa RH
- Subjects
- Animals, Base Sequence, DNA-Binding Proteins metabolism, Epithelium embryology, Epithelium metabolism, Hepatocyte Nuclear Factor 3-alpha, Humans, Kidney chemistry, Kidney Tubules, Collecting embryology, Kidney Tubules, Collecting metabolism, Liver chemistry, Liver cytology, Mice, Models, Biological, Molecular Sequence Data, NFI Transcription Factors, Rats, Rats, Sprague-Dawley, Thyroid Nuclear Factor 1, Transcription Factors genetics, Y-Box-Binding Protein 1, CCAAT-Enhancer-Binding Proteins, DNA-Binding Proteins genetics, Gene Expression Regulation, Nuclear Proteins genetics, Nuclear Proteins metabolism, Promoter Regions, Genetic genetics, Transcription Factors metabolism
- Abstract
The hepatocyte nuclear factor-3 alpha (HNF-3 alpha) and -3 beta proteins share homology in the winged helix/fork head DNA binding domain and regulate cell-specific transcription in hepatocytes and respiratory epithelium. In this study, we used transfection assays to demonstrate that the -520 nucleotides upstream of the rat HNF-3 alpha gene were sufficient for cell-specific expression. We identified binding sites for a liver and kidney-enriched nuclear factor and a kidney-enriched protein that recognizes two distinct promoter elements. We showed that the rat HNF-3 alpha promoter binds the HNF-3 protein isoforms, which may serve an auto- and/or cross-regulatory role. Furthermore, we showed that cotransfection of the thyroid transcription factor-1 expression vector enhanced HNF-3 alpha promoter activity. We discuss these results with respect to the transcriptional induction of the HNF-3 alpha gene in respiratory epithelium during embryogenesis. Because the HNF-3 alpha promoter region bound nuclear factors in kidney extracts, we used in situ hybridization to demonstrate that it was expressed in the urothelium of the renal pelvis in adult and embryonic kidney. We also report on a novel expression pattern of HNF-3 alpha in the epithelium of the urinary bladder, penile urethra, and the prostate gland, and show that its expression in the intestinal epithelium increases from the proximal duodenum to distal ileum. We also demonstrate that HNF-3 alpha is abundantly expressed in the colonic epithelium. Furthermore, we use the HNF-3 DNA binding consensus sequence to identify putative target genes in the renal pelvis and gut epithelium.
- Published
- 1997
91. Identification of a transthyretin enhancer site that selectively binds the hepatocyte nuclear factor-3 beta isoform.
- Author
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Samadani U, Qian X, and Costa RH
- Subjects
- Animals, Binding Sites, CCAAT-Enhancer-Binding Proteins, HeLa Cells, Hepatocyte Nuclear Factor 3-beta, Humans, Mice, Recombinant Fusion Proteins metabolism, Tumor Cells, Cultured, DNA-Binding Proteins metabolism, Enhancer Elements, Genetic, Nuclear Proteins metabolism, Prealbumin genetics, Transcription Factors metabolism
- Abstract
The upstream proximal region of the transthyretin (TTR) promoter and a distal enhancer are sufficient to drive liver-specific expression of the TTR gene, as demonstrated by experiments in transgenic mice. Previous analyses have characterized the binding of a number of liver-enriched transcription factors of the TTR promoter including hepatocyte nuclear factors one (HNF-1), HNF-4, and three distinct HNF-3 proteins (alpha, beta, and gamma), which are members of the winged helix (fork head) family. The TTR enhancer was shown to bind members of the CCAAT/enhancer binding protein (C/EBP) family at two distinct sites (TTR-2 and TTR-3), and an oligonucleotide containing the activation protein one (AP-1) binding sequence competed for recognition to a third enhancer site (TTR-1). In this study, we have carried out a detailed analysis of the transcription factors that recognize the TTR enhancer elements (TTR-1, TTR-2, and TTR-3 oligonucleotide sequences). Analysis of the TTR-1 site demonstrates that the putative AP-1 site in the TTR enhancer binds a ubiquitously expressed factor that is distinct from the AP-1 family of proteins. Next we demonstrate, via gel shift analysis, that the TTR-3 site is recognized by the C/EBP family in liver nuclear extracts. We also show that whereas the TTR-2 enhancer site is capable of binding recombinant C/EBP proteins, it does not bind C/EBP proteins from liver nuclear extracts. The TTR-2 site does, however, contain a variant HNF-3 recognition sequence that exclusively binds the HNF-3 beta isoform. Mutation of this HNF-3 beta-specific recognition sequence caused reductions in TTR enhancer activity. We had previously observed a 95% decrease in HNF-3 alpha expression and a 20% reduction in HNF-3 beta expression in acute phase livers, which correlated with a 60% decrease in TTR gene transcription. We propose that the HNF-3 beta-specific binding site in the TTR enhancer may play a role in maintaining TTR gene expression during the acute phase response in spite of the dramatic reduction in HNF-3 alpha protein levels.
- Published
- 1996
92. Cytokine regulation of the liver transcription factor hepatocyte nuclear factor-3 beta is mediated by the C/EBP family and interferon regulatory factor 1.
- Author
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Samadani U, Porcella A, Pani L, Johnson PF, Burch JB, Pine R, and Costa RH
- Subjects
- Base Sequence, CCAAT-Enhancer-Binding Proteins, Carcinoma, Hepatocellular, DNA-Binding Proteins genetics, G-Box Binding Factors, Gene Expression immunology, HeLa Cells immunology, Hepatocyte Nuclear Factor 3-beta, Humans, Interferon Regulatory Factor-1, Interleukin-1 pharmacology, Interleukin-6 pharmacology, Leucine Zippers genetics, Liver immunology, Liver metabolism, Molecular Sequence Data, Nuclear Proteins genetics, Promoter Regions, Genetic genetics, Cytokines physiology, DNA-Binding Proteins immunology, DNA-Binding Proteins pharmacology, Nuclear Proteins immunology, Nuclear Proteins pharmacology, Phosphoproteins pharmacology, Transcription Factors immunology
- Abstract
Three distinct hepatocyte nuclear factor-3 (HNF-3) proteins (alpha, beta, and gamma) regulate the transcription of numerous liver-enriched genes. The HNF-3 proteins bind DNA via a homologous winged helix motif common to a number of proteins known to be critical for determination events in embryogenesis. We have demonstrated previously that two binding sites in the -184 HNF-3 beta promoter are recognized by widely distributed factors and that there is also a critical autoregulatory site, we identified a binding site for a cell-specific factor, LF-H3 beta, that may function in restricting HNF-3 beta gene expression to hepatocytes. Our present study demonstrates that members of the C/EBP and proline and acidic amino acid-rich subfamilies of basic region leucine zipper transcription factors bind the LF-H3 beta site, and cotransfection of HepG2 cells shows that these factors are able to activate an HNF-3 beta promoter reporter construct. The LF-H3 beta-C/EBP binding sequence also confers HNF-3 beta promoter stimulation in response to interleukin (IL)-1 and IL-6. Upstream of this HNF-3 beta proximal promoter region, an IFN-stimulated response element core sequence (-231 to -210) was found that mediates transcriptional induction by IFN-gamma but not IFN-alpha. Gel mobility supershift assay demonstrates that an IFN-gamma-induced protein-DNA complex is disrupted by an antibody specific for interferon regulatory factor-1/interferon-stimulated gene factor-2. Consistent with this finding, we observed that IFN-gamma induction requires ongoing protein synthesis. Surprisingly, the effect of the three cytokines (IL-1, IL-6, and IFN-gamma) in combination as assayed by the same model is not synergistic. HNF-3beta joins the C/EBP family on the list of liver-enriched transcription factors, the expression of which is modulated by cytokines.
- Published
- 1995
93. Analysis of hepatocyte nuclear factor-3 beta protein domains required for transcriptional activation and nuclear targeting.
- Author
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Qian X and Costa RH
- Subjects
- Amino Acid Sequence, Animals, Base Sequence, Binding Sites genetics, Casein Kinases, Cell Nucleus metabolism, Conserved Sequence, DNA genetics, DNA metabolism, DNA Primers genetics, DNA-Binding Proteins chemistry, Hepatocyte Nuclear Factor 3-beta, Humans, Molecular Sequence Data, Mutagenesis, Nuclear Proteins chemistry, Phosphorylation, Proline genetics, Protein Kinases, Sequence Deletion, Sequence Homology, Amino Acid, beta-Galactosidase metabolism, DNA-Binding Proteins genetics, DNA-Binding Proteins metabolism, Nuclear Proteins genetics, Nuclear Proteins metabolism, Transcription Factors, Transcriptional Activation
- Abstract
Three distinct hepatocyte nuclear factor 3 (HNF-3) proteins (alpha, beta and gamma) regulate transcription of the transthyretin (TTR) and numerous other liver-specific genes. The HNF-3 proteins bind DNA via a homologous winged helix motif common to a number of developmental regulatory proteins including the Drosophila homeotic fork head (fkh) protein. The mammalian HNF-3/fkh family consists of at least thirty distinct members and is expressed in a variety of different cellular lineages. In addition to the winged helix motif, several HNF-3/fkh family members also share homology within transcriptional activation region II and III sequences. In the present study we further define the sequence boundaries of the HNF-3 beta N-terminal transcriptional activation domain to extend from amino acids 14 to 93 and include conserved region IV and V sequences. We also demonstrate that activity of the HNF-3 N-terminal domain was diminished by mutations which altered a putative alpha-helical structure located between amino acid residues 14 and 19. However, transcriptional activity was not affected by mutations which eliminated two conserved casein kinase I sites or increased the number of acidic amino acid residues in the N-terminal domain. Furthermore, we determined that the nuclear localization signal overlaps with the winged helix DNA-binding motif. These results suggest that conserved sequences within the winged helix motif of the HNF-3/fkh family may be involved not only in DNA recognition, but also in nuclear targeting.
- Published
- 1995
- Full Text
- View/download PDF
94. Murine chromosomal location of eight members of the hepatocyte nuclear factor 3/fork head winged helix family of transcription factors.
- Author
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Avraham KB, Fletcher C, Overdier DG, Clevidence DE, Lai E, Costa RH, Jenkins NA, and Copeland NG
- Subjects
- Amino Acid Sequence, Animals, Crosses, Genetic, Genes, Hepatocyte Nuclear Factor 3-alpha, Hepatocyte Nuclear Factor 3-beta, Hepatocyte Nuclear Factor 3-gamma, Mice, Inbred C57BL, Mice, Mutant Strains, Molecular Sequence Data, Muridae genetics, Sequence Alignment, Sequence Homology, Amino Acid, Transcription Factors classification, Chromosome Mapping, DNA-Binding Proteins genetics, Mice genetics, Nuclear Proteins genetics, Transcription Factors genetics
- Abstract
A 100-amino-acid DNA-binding motif, known as the winged helix, was first identified in the mammalian hepatocyte nuclear factor-3 (HNF-3) and Drosophila fork head family of transcription factors. Subsequently, more than 40 different genes that contain the winged helix motif have been identified. In the studies described here, we have determined the murine chromosomal location of eight members of this gene family, HFH-1, HFH-3, HFH-4, HFH-5, HFH-6, HFH-8, BF-1, and BF-2, by interspecific backcross analysis. These genes, designated HNF-3 fork head homolog 1 (Hfh1), Hfh3, Hfh4, Hfh5, Hfh6, Hfh8, Hfh9, and Hfh10, respectively, mapped to 6 different mouse autosomes and are thus well dispersed throughout the mouse genome. Based on this mapping information, we predict the chromosomal location of these genes in humans and discuss the potential of these genes as candidates for uncloned mouse mutations.
- Published
- 1995
- Full Text
- View/download PDF
95. Members of the HNF-3/forkhead family of transcription factors exhibit distinct cellular expression patterns in lung and regulate the surfactant protein B promoter.
- Author
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Clevidence DE, Overdier DG, Peterson RS, Porcella A, Ye H, Paulson KE, and Costa RH
- Subjects
- Amino Acid Sequence, Animals, Base Sequence, Bronchi metabolism, Epithelium metabolism, Forkhead Transcription Factors, Gene Library, In Situ Hybridization, Liver metabolism, Lung cytology, Male, Mice, Mice, Inbred BALB C, Molecular Sequence Data, Nuclear Proteins metabolism, Organ Specificity, Pulmonary Alveoli metabolism, Rats, Rats, Sprague-Dawley, Recombinant Fusion Proteins, Sequence Homology, Amino Acid, Transcription Factors metabolism, Aging metabolism, Embryonic and Fetal Development, Gene Expression, Gene Expression Regulation, Lung metabolism, Nuclear Proteins biosynthesis, Promoter Regions, Genetic, Proteolipids biosynthesis, Pulmonary Surfactants biosynthesis, Transcription Factors biosynthesis
- Abstract
The hepatocyte nuclear factor-3 (HNF-3)/forkhead (fkh) proteins consist of an extensive family of tissue-specific and developmental gene regulators which share homology within the winged helix DNA binding motif. We report on the isolation of a new family member, HNF-3/forkhead homolog 8 (HFH-8), from lung cDNA libraries and the derivation of the complete amino acid sequences for the HFH-8 protein as well as previously identified HFH-1 and HFH-4 proteins. The HFH proteins contain several sequence motifs found in activation domains of other transcription factors and HNF-3/fkh family members. In situ hybridization with the HNF-3, HFH-4, and HFH-8 probes in adult lung demonstrate that the HNF-3/fkh cellular expression patterns are regionally specified. Whereas HNF-3 alpha and HNF-3 beta are normally coexpressed in the hepatocyte, their expression patterns in the lung are different. The HNF-3 alpha and HFH-4 genes are coexpressed in the bronchiolar epithelium (clara cells), whereas the HNF-3 beta probe exhibits prominent hybridization with the smooth muscle surrounding arterioles and bronchioles. In contrast, HFH-8 probes labeled the type II pneumocyte cells lining the respiratory surfaces of terminal bronchioles and alveolar sac. We have identified an HNF-3 consensus DNA binding sequence in the proximal surfactant protein B (SPB) promoter region (SPB-f2, -78 to -88). SPB gene transcription is restricted to bronchiolar and alveolar epithelium which colocalizes with the expression pattern of the HNF-3 alpha and HFH-8 genes, respectively. We show that the SPB-f2 sequence is recognized by both HNF-3 alpha and HFH-8 proteins and that these cDNA expression vectors activate the SPB promoter in cotransfection assays through the HNF-3 consensus sequence. Our results suggest that SPB promoter activity is regulated by HNF-3 alpha and HFH-8 proteins in a cell type-specific manner.
- Published
- 1994
- Full Text
- View/download PDF
96. Hepatocyte nuclear factor-3 (HNF-3) binds to the insulin response sequence in the IGF binding protein-1 (IGFBP-1) promoter and enhances promoter function.
- Author
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Unterman TG, Fareeduddin A, Harris MA, Goswami RG, Porcella A, Costa RH, and Lacson RG
- Subjects
- 3T3 Cells, Animals, Base Sequence, Binding Sites, Cell Nucleus metabolism, Consensus Sequence, Hepatocyte Nuclear Factor 3-alpha, Hepatocyte Nuclear Factor 3-beta, Hepatocyte Nuclear Factor 3-gamma, Insulin pharmacology, Insulin-Like Growth Factor Binding Protein 1, Mice, Molecular Sequence Data, Mutagenesis, Site-Directed, Oligodeoxyribonucleotides, Recombinant Proteins metabolism, Recombinant Proteins pharmacology, Transfection, Carrier Proteins genetics, DNA-Binding Proteins metabolism, Insulin metabolism, Nuclear Proteins metabolism, Promoter Regions, Genetic, Transcription Factors metabolism
- Abstract
IGF binding protein-1 is an important short-term modulator of IGF bioavailability. Hepatic transcription of IGFBP-1 is increased by glucocorticoids and suppressed by insulin. We previously identified adjacent glucocorticoid and insulin response sequences approximately 90 bp 5' to the RNA cap site in the IGFBP-1 promoter. This insulin response sequence contains a sequence highly related (10/12 bases) to a consensus HNF-3 binding sequence. Gel shift and supershift studies confirm that this sequence binds HNF-3 alpha, beta and gamma. Co-expression of HNF-3 beta enhances IGFBP-1 promoter activity in NIH-3T3 cells. Mutation of this HNF-3 binding sequence disrupts this effect as well as the ability of glucocorticoids to stimulate and of insulin to inhibit IGFBP-1 promoter activity in H4IIE and HepG2 hepatoma cells. HNF-3 binding at this site may play an important role in the multihormonal regulation of hepatic IGFBP-1 gene expression.
- Published
- 1994
- Full Text
- View/download PDF
97. Retinoic acid-mediated activation of HNF-3 alpha during EC stem cell differentiation.
- Author
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Jacob A, Budhiraja S, Qian X, Clevidence D, Costa RH, and Reichel RR
- Subjects
- Animals, Base Sequence, Carcinoma, Embryonal, Cell Differentiation, DNA metabolism, DNA-Binding Proteins genetics, Gene Expression Regulation, Hepatocyte Nuclear Factor 3-alpha, Mice, Molecular Sequence Data, Nuclear Proteins genetics, Promoter Regions, Genetic, Protein Binding, RNA, Messenger analysis, Stem Cells cytology, Tumor Cells, Cultured, DNA-Binding Proteins metabolism, Nuclear Proteins metabolism, Stem Cells metabolism, Transcription Factors metabolism, Tretinoin metabolism
- Abstract
We present evidence demonstrating that the liver-enriched transcription factor HNF-3 alpha is activated upon retinoic acid-induced differentiation of mouse F9 embryonal carcinoma cells. We have detected increases in the DNA binding activity and mRNA level of HNF-3 alpha. Both are reflections of the actual activation mechanism at the level of transcriptional initiation, which we showed with the help of HNF-3 alpha promoter constructs. Time course studies clearly show that HNF-3 alpha activation is a transient event. Employing Northern blots, HNF-3 alpha mRNA can be detected between 16 and 24 hours post-differentiation, reaches its zenith at approximately 1 day, and then declines to virtually undetectable levels. F9 cells can give rise to three distinct differentiated cell types; visceral endoderm, parietal endoderm, and primitive endoderm. We have clearly shown that HNF-3 alpha stimulation occurs upon primitive endoderm formation. In addition, the transcription factor is also activated during the induction of cell lineages that give rise to parietal and visceral endoderm. HNF-3 alpha stimulation upon visceral endoderm differentiation is accompanied by the activation of HNF-3 target genes such as transthyretin, suggesting that HNF-3 alpha is involved in the developmental activation of this gene. In contrast, HNF-3 alpha target genes in parietal and primitive endoderm have yet to be identified. However, the stimulation of HNF-3 alpha during primitive endoderm formation, which is an extremely early event during murine embryogenesis, points towards a role for the factor in crucial determination processes that occur early during development.
- Published
- 1994
- Full Text
- View/download PDF
98. Angiotensin converting enzymes from urine of treated and untreated essential mild hypertensive patients (EHP) with diuretic: partial purification and characterization.
- Author
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Alves KB, Casarini DE, da Costa RH, Plavnik FL, Portela JE, and Marson O
- Subjects
- Adult, Blood Pressure, Chromatography, DEAE-Cellulose, Chromatography, Gel, Electrophoresis, Polyacrylamide Gel, Female, Humans, Hypertension urine, Kinetics, Male, Middle Aged, Molecular Weight, Peptidyl-Dipeptidase A isolation & purification, Reference Values, Time Factors, Chlorthalidone therapeutic use, Hypertension drug therapy, Hypertension enzymology, Peptidyl-Dipeptidase A urine
- Abstract
Urine of untreated EHP was eluted, on a ion-exchange chromatography, in two protein peaks with ACE activity, at 0.7 mS (BI) and 1.25 mS (BII), while urine of treated EHP, was eluted only in one peak with ACE activity (0.7 mS). BI (Mr, 88 kDa) and BII (Mr, 61 kDa) convert AI to AII, hydrolyze bradikinin, are inhibited by captopril, EDTA and metal ions.
- Published
- 1992
99. Inhibition of aminopeptidase activity by aromatic and other cyclic compounds.
- Author
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Alves KB, Costa RH, Aguilar MA, and Freitas Júnior JO
- Subjects
- Aminopeptidases drug effects, Aminopeptidases urine, Animals, Dose-Response Relationship, Drug, Humans, Kidney enzymology, Rats, Seeds enzymology, Glycine max enzymology, Substrate Specificity drug effects, Trees enzymology, Aminopeptidases antagonists & inhibitors, Polycyclic Compounds pharmacology
- Abstract
The effect of 2-naphthylamine, p-nitroaniline, o-phenanthroline, sodium deoxycholate and hydrocortisone succinate on the activity of human urine aminopeptidase, rat kidney methionyl and arginyl aminopeptidase, soybean and Enterolobium contortisiliquum seed aminopeptidase was studied using aminoacyl-2-naphthylamide and L-Leu-p-nitroanilide as substrates. Ki values ranged from 10 microM to 2.7 mM. On the basis of Ki and Km values, and catalytic efficiency for each enzyme, it is clear that the aminopeptidases from human urine and from soybean seed should be assayed with both substrates, whereas L-Leu-p-nitroaniline is a more appropriate substrate for the rat kidney aminopeptidases. Sodium deoxycholate is a better inhibitor than hydrocortisone succinate. Non-competitive inhibition was observed in all cases except for E. contortisiliquum seed aminopeptidase.
- Published
- 1992
100. Site-directed mutagenesis of hepatocyte nuclear factor (HNF) binding sites in the mouse transthyretin (TTR) promoter reveal synergistic interactions with its enhancer region.
- Author
-
Costa RH and Grayson DR
- Subjects
- Animals, Base Sequence, Basic Helix-Loop-Helix Leucine Zipper Transcription Factors, Binding Sites, DNA-Binding Proteins genetics, Gene Expression Regulation physiology, Hepatocyte Nuclear Factor 1, Hepatocyte Nuclear Factor 1-alpha, Hepatocyte Nuclear Factor 1-beta, Hepatocyte Nuclear Factor 3-alpha, Hepatocyte Nuclear Factor 3-beta, Hepatocyte Nuclear Factor 4, Humans, Mice, Models, Genetic, Molecular Sequence Data, Mutagenesis, Site-Directed physiology, Nuclear Proteins genetics, Nuclear Proteins metabolism, Transcription Factors genetics, Transfection genetics, Tumor Cells, Cultured, DNA-Binding Proteins metabolism, Enhancer Elements, Genetic physiology, Phosphoproteins, Prealbumin genetics, Promoter Regions, Genetic physiology, Transcription Factors metabolism
- Abstract
The transthyretin (TTR) gene is regulated by two DNA regions which elicit hepatocyte-specific expression: a proximal promoter and distal enhancer. The TTR promoter and enhancer are composed of at least eight DNA binding sites for three different hepatocyte nuclear factors (HNF), CCAAT/enhancer binding protein (C/EBP), and AP-1/cJun. Site directed mutations within each of the HNF binding sites in the TTR promoter were introduced to evaluate their contribution to transcriptional activity in hepatoma cells. The data indicate that the strong affinity HNF-3-S binding site (-106 to -94) is absolutely required for TTR promoter activity since several mutations in this site eliminate TTR expression in the context of its enhancer. Conversion of a second weak affinity HNF3-W site (-140 to -131) in the TTR promoter to a high affinity site resulted in higher levels of expression. TTR mutations that disrupted several weak affinity sites (HNF1, HNF3-W, and HNF4) only slightly diminished expression levels in the presence of the TTR enhancer. In contrast, when we deleted the TTR enhancer from these HNF mutant constructs, TTR expression decreased to undetectable levels. This result suggests cooperation between the factors binding to the TTR promoter and enhancer regions. These results also demonstrate that the HNF3-S site alone is not sufficient to activate TTR transcription, but rather requires the participation of three cell-specific factors to elicit minimal promoter activity. The complexity of this promoter design and the requirement for a minimal number of cell-specific factors to achieve transcription allows us to propose a model which may explain the maintenance of tissue-specific expression of TTR.
- Published
- 1991
- Full Text
- View/download PDF
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